Granulocyte Colony Stimulating Factor
Granulocyte Colony Stimulating Factor (G-CSF), in its natural form comprises two forms: a protein having 174 amino acids, and a form having three additional amino acids. Both forms have five cysteine residues; four forming two disulfide bonds, and one free. In its natural form G-CSF is a glycoprotein. G-CSF supports the growth of predominantly neutrophil colonies in a colony-forming (CFU-GM) assay, and in the presence of accessory cells, supports the growth of early erythroid (BFU-E) and pluripotential progenitors (CFU-GEMM) (granulocytes, erythrocytes, monocytes, and macrophages). G-CSF is also capable of promoting the differentiation of some myeloid leukemic cell lines (e.g., HL-60, WEHI-3B-D.sup.+), fresh myeloid leukemic cells, and has been reported to enhance the chemotactic peptide binding on peripheral blood neutrophils. In addition, G-CSF can significantly increase the ability of neutrophils to kill tumor targets in vitro through antibody-dependent cellular cytotoxicity (ADCC). In vivo experiments with recombinant human G-CSF in hamsters indicate a specific action on the neutrophil lineage with increases of three to sixfold in peripheral blood neutrophils.
Because of its hydrophobic characteristics, G-CSF is difficult to formulate. Examples of attempts to formulate G-CSF are shown in UK Ptent Application GB 2193631. Detergents, such as Tween-80, have been used to maintain G-CSF in a monomeric form and to minimize particulate formation. G-CSF contains five cysteinyl residues, four of which are involved in intrachain disulfide linkage. The free cysteinyl residue is, in general, inaccessible to modification due to folding of the molecule. At elevated temperatures the molecule "breaths" more rapidly and, on occasion, the cysteinyl residues interacts with each other leading to dimer and multimer formation. This phenomenon is enhanced by the presence of a non-ionic surfactant, e.g., Tween-80.TM. (polyoxyethylene sorbitan monooleate).